Fas is a cell surface antigen which transmits an apoptosis signal to the cell, and Fas is recognized by Fas antibody (Yonehara, S. et al., J. Exp. Med. vol. 169, 1747–1756, 1989), which is a monoclonal antibody produced by immunizing a mouse with human fibroblast. Fas gene was recently cloned by Itoh, N. et al., and it was then found out that Fas is a cell membrane protein of about 45 kD, and from the amino acid sequence, it was revealed that Fas is a member of the TNF receptor family (Cell, vol. 66, pages 233–243, 1991). The mouse Fas gene was also cloned (Watanabe-Fukunaga, R. et al., J. Immunol., vol. 148, pages 1274–1279, 1992), and its expression in thymus, liver, lung, heart, ovary was confirmed.
Human Fas ligand is a polypeptide which has been reported by Nagata et al. to be a biological molecule which induces apoptosis of Fas-expressing cells (Tomohiro Takahashi et al., International Immunology, vol. 6, pages 1567–1574, 1994). Human Fas ligand is a Type II membrane protein of the TNF family with a molecular weight of about 40 kD. As in the case of TNF, human Fas ligand in the human body is estimated to be in the form of a trimer (Masato Tanaka et al., EMBO Journal, vol. 14, pages 1129–1135, 1995). The extracellular domain of human Fas ligand is highly homologous with the extracellular domain of rat Fas ligand (Takashi Suda et al., Cell, vol. 75, pages 1169–1178, 1993) and mouse Fas ligand (Tomohiro Takahashi et al., Cell, vol. 76, pages 969–976, 1994). Human Fas ligand recognizes not only human Fas but also mouse Fas to induce apoptosis; and vice versa, rat Fas ligand and mouse Fas ligand also recognize human Fas to induce apoptosis.
Considerable researches have also been done on the mechanism of signal transduction in the cell upon Fas-mediated apoptosis, and identification and cloning of the factor which interacts with the intracellular domain of Fas, in particular, the region called “death domain” to transmit or block the signal has been reported. Possibility of the involvement of interleukin-1-converting enzyme (ICE)-related thiol proteases in Fas-mediated apoptosis has also been indicated.
A relationship between apoptosis, in particular, Fas-mediated apoptosis with various diseases and physiological phenomena has been recently indicated. For example, possibility has been indicated for involvement of abnormal Fas-mediated apoptosis in the decline of the T cell count in patients suffering from AIDS, in the death of hepatocytes in viral fulminant hepatitis, in some types of autoimmune diseases, and the like.
Involvement of the Fas/Fas ligand system in functions other than apoptosis has also been indicated. For example, the possibility has been indicated for the Fas/Fas ligand system to react with neutrophils to develop a proinflammatory action (Kayagaki, N. et al., Rinshou Meneki (Clinical Immunology), vol. 28, pages 667–675, 1996).
The condition wherein neutrophils and other immunocompetent cells are activated by various stimuli, such as endotoxins or by invasion, and humoral factors, such as cytokines are released into the blood and the tissues to induce systemic inflammatory responses is referred as systemic inflammatory response syndrome (hereinafter abbreviated as SIRS). SIRS is often associated with various organ failures, and when such organ failures are serious, SIRS may result in multiple organ failure syndrome (MODS) (Wakabayashi et al., Rinsho Kensa (Laboratory test), vol. 38, pages 349–352, 1994). Various factors are said to be involved in the process of organ failure, and the role in such organ failure of the invasion and accumulation of neutrophils by the action of IL-8 produced in macrophages and of other cells by IL-1 locally produced in response to invasion is of current interest. As described above, the Fas/Fas ligand system (hereinafter referred to as Fas/FasL system) is also reported for the possibility of its involvement in the activation of the neutrophils.
Ischemic reperfusion injury is found in practically all tissues and organs, and is involved in various diseases. Ischemic reperfusion injury is also a problem in preservation and transplantation of organs. Among such ischemic reperfusion injuries, those associated with infarction of liver, heart or kidney and those associated with surgery or transplantation, and in particular, tissue injury (such as cell necrosis) and dysfunction (such as cardiac arrhythmia) in the particular organ may lead to the death of the individual when they are serious, and therefore, such cases are a serious social problem. Various organ failures and ischemic reperfusion injuries from the early stage to the late stage are known to be associated with production and secretion of IL-8. It is also known that organ preservation and reperfusion in the course of organ transplantation is associated with the occurrence of the apoptosis. In addition, observation of apoptosis and fluctuation in the expression of Fas or FasL have been reported for some experimental models. There have also been reported a marked increase in the number of neutrophils at 24 hours after the reperfusion of liver after ischemia, and improvement of ischemic reperfusion injury by a neutralizing antibody of neutrophils (Jaeschke, H. et al., FASEB Journal, vol. 4, pages 3355–3359, 1990), and a marked increase of IL-8, neutrophils and macrophages at 3 hours after the reperfusion of the lung after ischemia, and improvement of ischemic reperfusion injury by a neutralizing antibody of the IL-8 (Sekido, N. et al., Nature, vol. 365, pages 654–657, 1993). These findings suggest significant roles of neutrophils and IL-8 in organ failures and ischemic reperfusion injuries of from the early stage to the late stage. On the other hand, it is not yet found how Fas/FasL is involved in such failures and injuries.
In infection by bacteria, endotoxin induces production of various cytokines in the body resulting in, for example, endotoxin shock in endotoxemia and sepsis as well as various organ damages including liver damage (Dinarello, C. A. et al., J. American Medical Association, vol. 269, page 1829, 1993), and serious conditions are more than often induced. Observation of apoptosis in such processes and the possibility of some involvement in such processes of Fas/FasL have been reported in experimental studies. However, it is not yet found how Fas/FasL is involved in such failures.
Death of cardiomyocytes in the case of a heart disease has been believed to occur mainly through necrosis. However, possibility of some involvement of apoptosis, and in particular, involvement of Fas-mediated apoptosis in such heart disease is now reported for clinical practices and in experiments. For example, it has been reported that the level of Fas expression increases when neonatal rat cardiomyocytes are placed in in vitro ischemic conditions (Tanaka, M. et al., Circ. Res., 75, 426–433, 1994); and that NO may have some relation to the remodeling of plaques in arteriosclerosis since IL-1 induces not only the synthesis of nitrogen monoxide (NO) of vascular smooth muscle cells but also apoptosis, apoptosis by IL-1 is inhibited by an inhibitor of NO synthesis, and Fas expression is induced by NO (Fukuo, K. et al., Hypertension, 27, 823–826, 1996). It has also been reported that apoptosis of cardiomyocytes is found in a canine heart failure model and such apoptosis is associated with an increased Fas expression (Lab. Invest., 73, 771–787, 1995), and that most of the cardiomyocyte death in canine the myocardial infarction model occurs through apoptosis, and such cardiomyocyte death is associated with a 100-fold increase in Fas expression (Lab. Invest., 74, 86–107, 1996). Furthermore, Fukuda et al. examined Fas expression in cardiomyocytes of cardiomyocardial disease patients, and found no Fas expression in hypertrophic cardiomyopathy but some Fas expression in at least a part of cardiomyocytes in cases of myocarditis and dilated cardiomyopathy (Idiopathic Cardiomyopathy Investigation Group, 1994 Business Year Research Report, 152–155, 1995). However, it is yet to be found out how Fas is involved in such heart diseases. Accordingly, the reports as described above have provided no data with regard to whether Fas acts to promote the apoptosis of the cardiomyocytes or to suppress the apoptosis of the cardiomyocytes in heart diseases, and it was still unclear whether Fas is directly involved in cytotoxicity or death of the cardiomyocytes of the patients suffering from heart diseases. As a consequence, no therapeutic agent and no therapy for heart disease wherein the disease, is treated by inhibiting the Fas-mediated apoptosis is known to date.
Involvement of apoptosis is also indicated for renal diseases, and increase in Fas mRNA expression is reported in an experimental model of renal ischemic reperfusion injury (Haruno, N. et al., Endocrinology, vol. 137, 1938–1948, 1996). However, it is yet to be found out how the Fas/FasL system is involved in such renal diseases.
Graft versus host disease (hereinafter referred to as GVHD) is a disease caused by graft versus host reaction (GVH reaction), which is an immunoreaction that may occur upon transplantation of lymphocytes of a donor or a graft, against the tissue antigens of the host. Exemplary GVHDs are GVHD after bone marrow transplantation, such as with incompatible bone marrow transplantation or with bone marrow transplantation in congenital immune deficiency syndrome; GVHD after organ transplantation; GVHD after blood transfusion, in which a large amount of blood is transfused to a patient of hypoimmunity; and the like. GVHD is associated with organ or tissue failure based on GVH reaction, and diarrhea, exhaustion such as weight loss and thinning, exanthem, splenomegaly, and liver dysfunction are clinically observed. GVHD is also associated with histological symptoms such as disorganization of bone marrow and lymphoid tissue and atrophy of intestinal villi.
Death of the cells constituting the host tissue in various GVHD has been believed to occur mainly through necrosis. However, the possibility of some involvement of apoptosis, and in particular, Fas-mediated apoptosis in such GVHD is now reported from experiments. For example, it has been reported that death of epithelial cells in intestine, skin and tongue in a mouse GVHD model occurs mainly through apoptosis (Aniti C. Gilliam et al., J. Invest. Dermatol., vol. 107, pages 377–383, 1996). With regard to the involvement of Fas-mediated apoptosis, it has been reported that no difference was found in survival time between cases when the donor was spleen lymphocytes from a control mouse with normal Fas ligand and cases when the donor was spleen lymphocytes from a gld mouse, which is a Fas ligand-mutated mouse, and practically no damage in skin and liver was induced (Matthew, B., Barker, B. et al., J. Exp. Med., vol. 183, 2645–2659, 1996). In spite of the indication of the involvement of Fas-mediated apoptosis in GVHD, there is no conclusion with regard to whether the Fas-mediated apoptosis is related to mortality with the GVHD. Furthermore, the report as described above utilizes spleen lymphocytes from a gld mouse for the material, and it is likely that the GVHD reaction is influenced by alteration in the amount of the expression of factors other than Fas ligand (such as perforin and TNF) as a substitute for the lack of Fas ligand, and the results obtained may not necessarily reflect the genuine effect of the lack of Fas ligand. Therefore, it is yet to be found out how Fas-mediated apoptosis is involved in GVHD, and whether a substance which specifically inhibits Fas-mediated apoptosis can be used as a therapeutic agent for GVHD.
Nonspecific immunosuppressives, such as cyclosporin, that have been used as a prophylactic or therapeutic agent of the GVHD generate nonspecific immunosuppression, and therefore, suffer from adverse side effects such as infections. No therapeutic agent and no therapy for GVHD wherein the GVHD is treated by inhibiting Fas-mediated apoptosis are known to date. In addition, no therapeutic agent and no therapy for GVHD wherein the GVHD is treated by utilizing selective immunosuppression are known to date.
With regard to diseases based on ischemic reperfusion injury, commercially available drugs mainly aim at thrombolysis and improvement of circulation, and no drug is available that directly prevents or treats the damage. With regard to the endotoxemia and sepsis, steroid and proteolytic enzyme inhibitor, for example, are used in the case of shock, and no drug is currently available that directly prevents or treats the organ damage. The drugs used for the diseases based on organ damage mainly aim at palliative treatment, and no drug is available that prevents or radically treats the diseases based on organ damage. In addition, no prophylactic or therapeutic agent which is widely effective for various tissues and organs is available.
In view of such situation, there is a demand for a pharmaceutical which is effective in preventing or treating diseases based on damage of the tissue or the organ in a wide variety of tissues or organs, which is effective in vivo, and which is less toxic to humans. However, no pharmaceutical is so far available that meets such requirements.
An object of the present invention is to provide a pharmaceutical and a therapy in the form of a prophylactic/therapeutic agent or an organ preservative agent which contains a Fas antagonist and which works through a new mechanism. More particularly, the present invention provides a prophylactic/therapeutic agent, an organ preservative agent, which contains a Fas antagonist, and a therapy, in which a Fas antagonist are used for diseases wherein Fas is involved.